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David. J Cancer Metastasis Treat 2022;8:32 https://dx.doi.org/10.20517/2394-4722.2022.71 Page 7 of 12

[49]
deletion of Hnf4a in a mouse model of PDA accelerated tumorigenesis . Basal-like and classical PDA are
metabolically distinct, with basal-like tumors displaying a higher degree of glycolysis and classical tumors
[50]
exhibiting a more lipogenic phenotype . HNF4A depletion in PDA cells promoted increased glycolysis,
[44]
presumably mediated by increased expression of key enzymes in the glycolytic pathway . It is unclear
whether HNF4A directly regulates the expression of glycolytic enzymes, or whether the metabolic
phenotype upon HNF4A knockdown is the indirect result of activation of oncogenic pathways such as
PI3K, which were also activated by HNF4A depletion. In any event, despite their shared role in maintaining
classical gene expression, GATA6 and HNF4A appear to control distinct sets of genes, and the glycolytic
phenotype upon depletion was specific to HNF4A . Like HNF4A and GATA6, HNF1A is another
[44]
endodermal TF whose expression is associated with the classical subtype of PDA. As with the other TFs
associated with the classical subtype, pancreas-specific KO of Hnf1a coupled with mutant KRAS activation
accelerated tumorigenesis, and the emergence of tumors with sarcomatoid features . The same study also
[51]
showed that HNF1A cooperates with KDM6A to promote acinar cell differentiation, consistent with the fact
that knockout of either factor in acinar cells results in widespread ADM. Contrary to these results in
autochthonous mouse models, HNF1A has also been shown in some contexts to promote tumorigenesis.
Using primary human PDA cell lines, HNF1A was found to be a pro-tumor factor expressed more highly in
high
high
[52]
an EPCAM /CD44 population enriched with tumor-forming ability . These authors went on to show
that in the setting of human cell lines, HNF1A depletion inhibits tumor formation. It is currently unclear
what determines whether HNF1A plays a pro- or anti-tumor role in PDA cells.
Another series of studies performed in human cell lines aimed at examining the transcriptional
determinants of PDA tumor grade, which the authors noted is closely linked with classical (low grade) and
basal-like (high grade) subtypes [53,54] . The authors first assembled a collection of cell lines that represented
low- and high-grade tumors, and performed epigenomic profiling, identifying sets of enhancers marked by
H3K27 acetylation specific to each grade. This approach was combined with RNA-seq to identify TFs
differentially expressed in cell lines of each grade. The most strongly enriched motif in enhancers specific to
the low-grade cells was that of the ETS-domain family of TFs, consistent with the fact that family member
ELF3 was the second most highly expressed TF in low-grade compared to high grade. KLF5 was the most
highly expressed TF in low-grade versus high-grade tumors, and despite its motif not showing enrichment
in low-grade enhancers, the authors demonstrated that KLF5 is an important determinant of epithelial
differentiation in low-grade tumor cell lines . Unlike other TFs linked with the classical subtype, KLF5
[53]
knockout in mouse models completely abolished tumorigenesis, apparently by preventing acinar-to-ductal
metaplasia, an essential first step in tumorigenesis initiated in acinar cells [10,55] .

While most of the classical subtype-linked TFs described so far have been endoderm-derived lineage
regulators, another low-grade-specific TF identified in these studies is interferon regulatory factor 1 (IRF1),
[56]
a broadly expressed immune regulatory TF important for the interferon response . IRF1 is activated by
KLF5 and ELF3, explaining its expression in low-grade tumor cells. While IRF1 was not shown to play a
role in maintaining classical subtype identity, its expression in low-grade tumor cells resulted in greater
interferon responsiveness, resulting in higher expression of antigen presentation and processing genes,
echoing observations made in experiments targeting GATA6 [45,56] . Contrary to classical tumors, basal-like
tumors upregulate ZBED2, a zinc finger protein that binds to the promoters of IFN-regulated genes and
antagonizes their activation by IRF1 . These observations regarding differential IFN responsiveness of the
[57]
two subtypes appear consistent with the observation that basal-like PDA tumors are depleted of infiltrating
T cells relative to other subtypes , and again suggest that tumor subtype may play a role in determining
[24]
sensitivity to immunotherapy.